Effects of overexpression of glutamine:fructose-6-phosphate amidotransferase (GFAT) and glucosamine treatment on translocation of GLUT4 in rat adipose cells.
Insulin resistance is associated with diabetes. Hyperglycemia per se causes insulin resistance as well as increased flux of glucose through the hexosamine biosynthetic pathway. The rate-limiting enzyme for entry of glucose into this pathway is glutamine:fructose-6-phosphate amidotransferase (GFAT). To directly evaluate the role of GFAT in modulating insulin-stimulated glucose transport, we co-transfected primary cultures of rat adipose cells with expression vectors for human GFAT as well as an epitope-tagged GLUT4 and examined the effect of overexpressed GFAT on insulin-stimulated translocation of GLUT4. When we measured cell surface tagged GLUT4 in response to insulin, cells overexpressing GFAT and tagged GLUT4 had an insulin-dose response curve that was similar to that of control cells expressing only tagged GLUT4. As an alternative means of increasing flux through the hexosamine biosynthetic pathway, we incubated adipose cells with glucosamine (a substrate of the pathway downstream from GFAT) and insulin. Interestingly, for short incubation times (4 h) we observed a decrease in both basal and insulin-stimulated glucose transport without a detectable effect on insulin-stimulated translocation of GLUT4. However, for longer incubation times (16 h), we observed a significant decrease in the amount of GLUT4 in the plasma membrane. Our data suggest that products of the hexosamine biosynthetic pathway may cause insulin resistance, in part, by acutely decreasing intrinsic activity of GLUT4 as well as chronically altering the amount of GLUT4 at the cell surface.
Chen
H
H
Hypertension-Endocrine Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892-1754, USA.
Ing
B L
BL
Robinson
K A
KA
Feagin
A C
AC
Buse
M G
MG
Quon
M J
MJ
D017667:Adipocytes / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D002478:Cells, Cultured; D015870:Gene Expression; D005944:Glucosamine / Q000494:pharmacology*; D051275:Glucose Transporter Type 4; D005945:Glutamine-Fructose-6-Phosphate Transaminase (Isomerizing) / Q000235:genetics* / Q000378:metabolism; D006595:Hexosamines / Q000096:biosynthesis; D006801:Humans; D007328:Insulin / Q000494:pharmacology; D007333:Insulin Resistance; D008297:Male; D009004:Monosaccharide Transport Proteins / Q000378:metabolism*; D009124:Muscle Proteins; D051381:Rats; D014162:Transfection
D051275:Glucose Transporter Type 4; D006595:Hexosamines; D007328:Insulin; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; C495692:SLC2A4 protein, human; C495694:Slc2a4 protein, rat; D005945:Glutamine-Fructose-6-Phosphate Transaminase (Isomerizing); D005944:Glucosamine